Complement activation by human IgG antibodies to galactose‐α‐1,3‐galactose

Abstract

Summary Some human antibodies may paradoxically inhibit complement activation on bacteria and enhance pathogen survival in humans. This property was also claimed for IgG antibodies reacting with terminal galactose‐α‐1,3‐galactose (Galα3Gal; IgG anti‐αGal), a naturally occurring and abundant antibody in human plasma that targets numerous different pathogens. To reinvestigate these effects, we used IgG anti‐αGal affinity isolated from a pool of normal human IgG and human hypogammaglobulinaemia serum as a complement source. Flow cytometry was performed to examine antibody binding and complement deposition on pig erythrocytes, Escherichia coli O86 and Streptococcus pneumoniae serotype 9V. Specific nanobodies were used to block the effect of single complement factors and to delineate the complement pathways involved. IgG anti‐αGal was capable of activating the classical complement pathway on all the tested target cells. The degree of activation was exponentially related to the density of bound antibody on E. coli O86 and pig erythrocytes, but more linearly on S. pneumoniae 9V. The alternative pathway of complement amplified complement deposition. Deposited C3 fragments covered the activating IgG anti‐αGal, obstructing its detection and highlighting this as a likely general caveat in studies of antibody density and complement deposition. The inherent capacity for complement activation by the purified carbohydrate reactive IgG anti‐αGal was similar to that of normal human IgG. We propose that the previously reported complement inhibition by IgG anti‐αGal relates to suboptimal assay configurations, in contrast to the complement activating property of the antibodies demonstrated in this paper.

The human natural anti‐αGal antibody targets common pathogens by broad‐spectrum polyreactivity

“…In general, phagocytosis of pneumococci requires opsonization by antibodies and complement fragments, especially fragments of C3. We recently reported that anti‐αGal bound on serotype 9V pneumococci activated the classical pathway of complement 38 . Here, we examine the opsonic effect of anti‐αGal bound on this pneumococcal serotype.…”

“…Structures of most capsular polysaccharides are reported, and none of these contains terminal Galα3Gal 48 . As antibody source, we used a well‐characterized and highly purified preparation of human anti‐αGal antibody of the IgG class 37,38 and examined its reactivity to 91 serotypes of S. pneumoniae by flow cytometry. A positive reaction was defined as more binding signal than the maximal signal observed with a control antibody of irrelevant specificity (IgG anti‐CD20 antibody).…”

“…Human hypogammaglobulinaemia serum was separated from whole blood by centrifugation. The blood was obtained from a person with hypogammaglobulinaemia (low IgG (<0·8 g/L), low IgA (<0·16 g/L) and low IgM (0·10 g/L)) but with normal activity of the complement system 38 . Blood cells were prepared from samples of 4 ml heparin‐stabilized freshly drawn blood from random blood donors.…”

“…Such broad pathogen reactivity likely involves polyreactivity. We recently demonstrated that anti‐αGal reacts with pneumococci of serotype 9V, 38 which does not contain terminal Galα3Gal in its capsule polysaccharide 39 . Broad‐spectrum bacterial reactivity is not unique to the anti‐αGal antibody among the naturally occurring antibodies.…”

Nanobodies Provide Insight into the Molecular Mechanisms of the Complement Cascade and Offer New Therapeutic Strategies

“…Jensen et al recently utilized hC3Nb1, hC3Nb2, and C1qNb75 to attribute the complement pathways elicited by these antibodies targeting the galactose-α-1,3-galactose carbohydrate. The authors reported that the anti-αGal antibodies elicit complement deposition by activating the classical pathway [ 121 ]. This basic research application is an example of how complement inhibitory nanobodies can be used to assign the contribution of the individual pathways to complement activation.…”